The homeotic genes of Drosophila dictate segmental identity. Their expression boundaries are fixed by members of the Polycomb group family of genes (PcG). The broad objective of this proposal is to determine the mechanisms of this repression. Understanding the control of differentiation in this model eukaryotic system is expected to aid greatly in understanding the pathological deregulation of differentiation and proliferation that occurs in cancer, birth defects and neurodegenerative diseases. The specific aims of this proposal are twofold. 1) The roles of individual PcG members in repression will be investigated using lexA- PcG fusion proteins to assay repressor activity toward reporter genes in transgenic flies. The biological significance of repression, and the sequence of events leading to formation of the "repressor complex" will be investigated using mutant flies lacking known PcG members. Repression of different types of genes will be assessed by moving the lexA anchoring sites to different chromosomal locations. 2) New members of the PcG will be sought using a novel F(1) screen, in which mitotic recombination between engineered FRT sites will be induced on homologous chromosomes, to produce mosaicism for distal PcG mutant genes. Mutations on the FRT chromosomes will be generated by standard methods. Candidate PcG genes showing strong homeotic phenotypes will be cloned by P-element excision, and sequenced.